Formation of a blood clot (thrombus) in a blood vessel entails two principal events: platelet aggregation and deposition of fibrin. Within seconds of vessel injury, resting platelets become activated and are bound to the exposed surface of the injured area by a phenomenon called platelet adhesion. Activation of platelets also leads to enhanced binding of the surface adhesion molecule gpIIbIIIa (CD41), causing platelets to bind to each other in a process called platelet aggregation to form a platelet plug. Inhibitors of platelet activation, or of gpIIbIIIa activity, have been shown to prevent platelet aggregation and, hence, to reduce thrombus formation. The platelet plug can stop bleeding quickly, but it must be reinforced by fibrin for long-term effectiveness, until the vessel injury can be permanently repaired.
The coagulation system, involved in thrombus formation, has a natural counterpart in the fibrinolytic system. In the process of blood coagulation, a cascade of enzyme activities are involved in generating a fibrin network which forms the framework of a clot. Degradation of the fibrin network (fibrinolysis) is accomplished by the action of the enzyme plasmin. Plasminogen is the inactive precursor of plasmin and conversion of plasminogen to plasmin is accomplished by cleavage of the peptide bond between arginine 561 and valine 562 of plasminogen. Under physiological conditions this cleavage is catalysed by tissue-type plasminogen activator (tPA) or by urokinase-type plasminogen activator (uPA).
If the balance between the clotting and fibrinolytic systems becomes locally disturbed, intravascular clots may form at inappropriate locations leading to conditions such as coronary thrombosis and myocardial infarction, deep vein thrombosis, stroke, peripheral arterial occlusion and embolism. In such cases, the administration of fibrinolytic agents has been shown to be a beneficial therapy for the promotion of clot dissolution.
Fibrinolytic therapy has become relatively widespread with the availability of a number of plasminogen activators such as tPA, uPA, streptokinase and the anisoylated plasminogen streptokinase activator complex, APSAC. Each of these agents has been shown to promote clot lysis, but all have deficiencies in their activity profile which makes them less than ideal as therapeutic agents for the treatment of thrombosis (reviewed by Marder and Sherry, New England Journal of Medicine 1989, 318: 1513-1520). One of the major problems with tPA for the treatment of acute myocardial infarction or other thrombotic disorders is that it is rapidly cleared from the circulation with a plasma half-life in man of around 5 minutes (Bounameaux et al. in: "Contemporary Issues in Haemostasis and Thrombosis" vol 1 pp. 5-91, 1985. Collen et al. eds, Churchill Livingstone). This results in the need to administer tPA by infusion in large doses. The treatment is therefore expensive and is usually delayed since the patient has to be hospitalized before treatment can commence. Urokinase, in either the single chain form or the two chain form, has a similar rapid plasma clearance and also requires administration by continuous infusion.
A major problem shared by all of these agents is that at clinically useful doses, they are not thrombus specific as they activate plasminogen in the general circulation. The principal consequence of this is that proteins such as fibrinogen involved in blood clotting are destroyed and dangerous bleeding can occur. This also occurs with tPA despite the fact that, at physiological concentrations, it binds to fibrin and shows fibrin selective plasminogen activation. Significant efforts have been expended to find mutant or other forms of tPA and other fibrinolytic or thrombolytic agents with desirable activity, specificity, and duration.
Accordingly, a need exists in the art for additional agents which can enhance fibrinolysis. The subject invention fills this and other needs.